Dob1p (Mtr4p) is a putative ATP-dependent RNA helicase required for the 3' end formation of 5.8S rRNA in Saccharomyces cerevisiae.
The temperature-sensitive mutation, dob1-1, was identified in a screen for dependence on overexpression of the yeast translation initiation factor eIF4B (Tif3p). Dob1p is an essential putative ATP-dependent RNA helicase. Polysome analyses revealed an under accumulation of 60S ribosomal subunits in the dob1-1 mutant. Pulse-chase labelling of pre-rRNA showed that this was due to a defect in the synthesis of the 5.8S and 25S rRNAs. Northern and primer extension analyses in the dob1-1 mutant, or in a strain genetically depleted of Dob1p, revealed a specific inhibition of the 3' processing of the 5.8S rRNA from its 7S precursor. This processing recently has been attributed to the activity of the exosome, a complex of 3'-->5' exonucleases that includes Rrp4p. In vivo depletion of Dob1p also inhibits degradation of the 5' external transcribed spacer region of the pre-rRNA. A similar phenotype was observed in rrp4 mutant strains and, moreover, the dob1-1 and rrp4-1 mutations show a strong synergistic growth inhibition. We propose that Dob1p functions as a cofactor for the exosome complex that unwinds secondary structures in the pre-rRNA that otherwise block the progression of the 3'-->5' exonucleases.
de la Cruz
J
J
Département de Biochimie Médicale, Centre Médical Universitaire, Université de Genève, Switzerland. Jesus.Cruz@medecine.unige.ch
Kressler
D
D
Tollervey
D
D
Linder
P
P
D003001:Cloning, Molecular; D053487:DEAD-box RNA Helicases; D039642:Eukaryotic Initiation Factors; D005095:Exoribonucleases / Q000502:physiology; D005656:Fungal Proteins / Q000235:genetics / Q000502:physiology; D005690:Galactose / Q000235:genetics; D016296:Mutagenesis; D009687:Nuclear Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D010448:Peptide Initiation Factors / Q000502:physiology; D010641:Phenotype; D020365:RNA Helicases; D012316:RNA Nucleotidyltransferases / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D012341:RNA, Ribosomal, 5S / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D012269:Ribosomal Proteins / Q000172:deficiency / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins
D039642:Eukaryotic Initiation Factors; D005656:Fungal Proteins; D009687:Nuclear Proteins; D010448:Peptide Initiation Factors; D012322:RNA Precursors; D012341:RNA, Ribosomal, 5S; D016601:RNA-Binding Proteins; C098788:RRP4 protein, S cerevisiae; D012269:Ribosomal Proteins; D029701:Saccharomyces cerevisiae Proteins; C083432:TIF3 protein, S cerevisiae; D012316:RNA Nucleotidyltransferases; D005095:Exoribonucleases; C101004:MTR4 protein, S cerevisiae; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases; D005690:Galactose